New species form when diverging populations become reproductively isolated. One cause of this isolation is hybrid incompatibility, the sterilit and lethality of interspecific hybrids. This proposal investigates two previously identified genes that interact to cause lethality in hybrids between the fruit fly species Drosophila melanogaster and D. simulans. Sequence analysis of both genes shows that they have evolved under adaptive evolution, a pattern also observed with other hybrid incompatibility genes. Identifying the specific selective forces causing this pattern of molecular evolution will therefore help elucidate the biological causes of speciation. This proposal is based on the recent discovery that these Drosophila hybrid incompatibility genes are required to repress transposable elements, selfish DNAs that can mobilize, reach high copy number, and cause significant damage to eukaryotic genomes. A range of genetic and molecular assays are proposed here to identify how these genes function in repressing transposable elements, including profiling the genome-wide pattern of transposition in mutant lines. Other phenotypic assays will investigate how these genes affect fertility and meiotic performance within their own host species. Attention will then turn to an integrated effort to determine how sequence divergence and adaptive evolution between species has altered specific functions of these genes. Transposable elements are a major cause of spontaneous mutation and DNA damage in eukaryotes including humans. The proposed studies will provide a high resolution view of how transposable element load contributes to host gene evolution and reproductive isolation between species.